Polypeptides With Enhanced Anti-Inflammatory And Decreased Cytotoxic Properties And Relating Methods

Abstract

The invention provides a polypeptide containing at least one IgG Fc region region, said polypeptide having a higher anti-inflammatory activity and a lower cytotoxic activity as compared to an unpurified antibody and methods of production of such polypeptide.

1 .- 22 . (canceled) 
     
     
         23 . A method of preparing a pharmaceutical preparation having anti-inflammatory activity, comprising:
 expressing, in a host cell, a recombinant polypeptide containing an IgG Fc region;   subjecting the expressed recombinant polypeptide to one or more purification steps to produce a preparation comprising the recombinant polypeptide;   treating the composition comprising the recombinant polypeptide with an alpha-(2,6) sialylatransferase and a donor of sialic acid to increase the fraction of recombinant polypeptides in the composition having alpha-2,6-linked sialic acid on N-glycans of the IgG Fc region thereby preparing a composition enriched for sialylated recombinant polypeptides, and   processing the composition enriched for sialylated recombinant polypeptides to produce a pharmaceutical preparation, wherein the processing comprises combining the recombinant polypeptides with a physiologically acceptable carrier, excipient or stabilizer, thereby preparing a pharmaceutical preparation having anti-inflammatory activity.   
     
     
         24 . The method of  claim 23  wherein the recombinant polypeptide does not comprise a Fab region. 
     
     
         25 . The method of  claim 23  wherein the recombinant polypeptide comprise a Fab region. 
     
     
         26 . The method of  claim 23  wherein the recombinant polypeptide is an antibody. 
     
     
         27 . The method of  claim 23  wherein the recombinant polypeptide is a monoclonal antibody. 
     
     
         28 . The method of  claim 26  wherein the antibody is chimeric antibody. 
     
     
         29 . The method of  claim 26  wherein the antibody is a humanized antibody. 
     
     
         30 . The method of  claim 23  further comprising subjecting the composition enriched for sialylated recombinant polypeptides to affinity chromatography to produce a composition further enriched for sialylated recombinant polypeptides. 
     
     
         31 . The method of  claim 23  where in the host cell is a mammalian cell line 
     
     
         32 . The method of  claim 31  wherein the mammalian cell line is a CHO cell line. 
     
     
         33 . The method of  claim 23  further comprising measuring the sialic acid content of the composition enriched for sialylated recombinant polypeptides. 
     
     
         34 . The method of  claim 32  wherein the step of measuring comprises one or more of: affinity chromatography, HPLC, and mass spectrometry. 
     
     
         35 . The method of  claim 23  further comprising measuring the anti-inflammatory activity of the composition enriched for sialylated recombinant polypeptides. 
     
     
         36 . The method of  claim 23  further comprising measuring the antibody dependent cell cytotoxicity of the composition enriched for sialylated recombinant polypeptides. 
     
     
         37 . The method of  claim 23  wherein the purification step comprises ion-exchange chromatography or affinity chromatography. 
     
     
         38 . The method of  claim 23  wherein the composition enriched for sialylated recombinant polypeptides exhibits an increased protective effect in a K/B×N serum-induced arthritis mouse model relative to the un-enriched composition. 
     
     
         39 . The method of  claim 23  wherein the host cell harbors a recombinant sialyltransferase. 
     
     
         40 . A pharmaceutical preparation made by the method of  claim 23 .